What does TFBSPred do?
TFBSPred predicts transcription factor binding sites in a selected genomic locus in specific cell types.

How do I cite TFBSPred?
If you used TFBSPred, please cite:
Zogopoulos, V. L., Spaho, K., Ntouka, C., Lappas, G. A., Kyranis, I., Bagos, P., Spandidos D. A., and Michalopoulos, I. (2021). TFBSPred: A functional transcription factor binding site prediction webtool for humans and mice. Int J Epigenetics 1, 9.

Which species are available?
You can search for human and mouse genes.

Why only mouse and not more species?
Mouse genome has around 90% degree of similarity with the human genome. Furthermore, there were DNaseI Hypersensitivity data available only for human and mouse.

Where do stored data come from?
The database contains:

• Pairwise alignments between Homo sapiens and Mus musculus genomes from the Ensembl Compara from Ensembl release 90
• DNaseI Hypersensitivity data from ENCODE at UCSC
• TFFM data from JASPAR
• Homo sapiens and Mus musculus gene data from Ensembl BioMart

Why is TFBSPred better than similar webtools?
TFBSPred takes advantage of various parameters. First of all, we are using the novel TFFM algorithm to predict TFBSs. We utilize the open chromatin regions in multiple different cells discovered by DNAse HS data and we intersect them with the respective conserved genomic region between human and mouse. Thus, we improve our chances of finding a biologically active region and reduce the chance of false positives. In addition, instead of specifying a fixed length of TFBS search upstream or downstream of the promoter, TFBSPred calculates the conserved open chromatin region on its own.

Why didn't autocomplete find my gene when I typed it?
You were probably using an alias for your gene. Only HGNC/MGI gene symbols are accepted.

Where can I find the HGNC/MGI symbol of my gene?
For humans you can use the GeneCards (recommended) or the HGNC website. For mouse you can use the MGI website.

How do I use a genomic position as input?
You have to visit Ensembl release 90 in order to identify your genomic position. Format the input as: Chromosome:Genomic-Location(Orientation) e.g. 4:102501329(+)

I’ve submitted my gene of interest and I’m redirected to a page with a table containing my gene, its available transcripts and their TSSs. What do I do next?
In order to proceed with the web tool, you must click on one of the available TSS. The Gene symbol, Ensembl Gene and Transcript stable ID tabs contain links to the HGNC/MGI and Ensembl websites.

Why are some of the TSS cells empty?
Multiple different transcripts may use the same TSS. In that case we omit writing this TSS more than once.

I’ve picked my TSS. What is next?
Please select at least 1 option of each category (Lineage, Tissue, Karyotype and Sex) or there will not be any results. Afterwards, the filtered cell types are presented.

Why do I have to select specific cells?
We recommend choosing those cells similar to what you are interested in. Different cell types with different treatments contain various open chromatin regions.

What is that TFFM threshold field?
It is the threshold value for the TFFM search. The range of acceptable values is from 0.85 to 1. The higher the threshold, the stricter will the TFBS search be. We recommend values around 0.9-0.95.

How long do I have to wait for the results?
On average, results will be presented in about a minute. Please do not resubmit the data until then.

I have submitted my choices and after about a minute I got a blank page! What went wrong?
There are two possible explanations. You might have chosen a genomic position which is not in a conserved region between human and mouse. Alternatively, it is possible that the cell lines picked might not have an open chromatin landscape in that position. In order to determine which of the two is the cause, we recommend selecting all the cell lines. If a result is not produced, that could be an indication that the gene is not expressed in the specific cell lines.

I have submitted my choices and after about a minute I got some results! What am I seeing?
On the results page, the first thing visible is the pairwise alignment of the open chromatin human and mouse conserved region dictated by the TSS and the cells chosen in Pair format. The organism chosen is always first in the pair.

Why is that position in a red background?
The TSS is highlighted red and the arrow above it is the orientation of transcription.

What does the View MultiFASTA link do?
It produces the pairwise alignment shown below as a MultiFASTA format sequence. It can be used as input to multiple alignment visualization programs such as Jalview or ClustalX for further analysis.

What do the numbers at the side of each line of the alignment mean?
This is the genomic position of the beginning and the end of each of the subsequences. This way the users can easily search for a part of the sequence in a genome browser. Each line has a length of 50 chars.

So, where are the TFBS predictions?
The TFBS results are located above each alignment block. Detailed results are located below the pairwise alignment of the conserved open chromatin region, and can be displayed by clicking the respective button. We only present the conserved TFBSs and their DNA target sequences are also in Pair format. The detailed results are in alphabetical order by the name of each TF, along with a link to the relevant JASPAR webpage. More than one TFBSs for a single TF may be discovered.

I found the TFBS of a TF I am interested in but I cannot find the target sequence on the conserved open chromatin alignment at the start.
The TFBS results take into account the strand where the target sequence is discovered. Try converting it to its reverse complementary sequence and then searching for it. If you still cannot find it, the sequence might be fragmented between 2 different lines or there are gaps in it. In this case, try searching for it using the position numbers.